Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction of the Ministry of Education, Huazhong Agricultural University, Wuhan 430070, China.
Key Laboratory of Swine Genetics and Breeding of the Ministry of Agriculture, Huazhong Agricultural University, Wuhan 430070, China.
Int J Mol Sci. 2023 Apr 5;24(7):6773. doi: 10.3390/ijms24076773.
Myogenic differentiation is a complex biological process that is regulated by multiple factors, among which long noncoding RNAs (lncRNAs) play an essential role. However, in-depth studies on the regulatory mechanisms of long noncoding RNAs (lncRNAs) in myogenic differentiation are limited. In this study, we characterized the role of the novel lncRNA , which is upregulated during porcine skeletal muscle satellite cell (PSC) differentiation in myogenesis. We found that affected the proliferation and differentiation of PSC in vitro. We performed quantitative polymerase chain reaction (qPCR), 5-ethynyl-20-deoxyuridine (EdU), western blotting, immunofluorescence staining, pull-down assays, and cleavage under targets and tagmentation (CUT and Tag) assays to clarify the effects and action mechanisms of . LncRNA inhibited myoblast proliferation based on analyses of cell survival rates during PSC proliferation. Functional analyses revealed that promotes cell differentiation and enhances myogenin (), myosin heavy chain (), and myocyte enhancer factor 2 () during myoblast differentiation. As determined by pull-down and RNA immunoprecipitation (RIP) assays, the lncRNA interacted with PBX/Knotted Homeobox 2 (PKNOX2). CUT and Tag assays showed that PKNOX2 was significantly enriched on the promoter after lncRNA knockdown. Our findings demonstrate that the interaction between lncRNA and PKNOX2 relieves the inhibitory effect of PKNOX2 on the promoter, increases its expression, and promotes PSC differentiation. This novel role of lncRNA sheds light on the molecular mechanisms by which lncRNAs regulate porcine myogenesis.
肌发生是一个复杂的生物学过程，受多种因素调控，其中长链非编码 RNA（lncRNA）起着重要作用。然而，lncRNA 在肌发生中的调控机制的深入研究还很有限。在这项研究中，我们描述了 novel lncRNA 在猪骨骼肌卫星细胞（PSC）分化中的作用，该 lncRNA 在肌生成过程中上调。我们发现 影响 PSC 在体外的增殖和分化。我们进行了定量聚合酶链反应（qPCR）、5-乙炔基-20-脱氧尿苷（EdU）、western blot、免疫荧光染色、下拉实验和切割靶标和标签（CUT 和 Tag）实验，以阐明 的作用和作用机制。基于 PSC 增殖过程中细胞存活率的分析，lncRNA 抑制成肌细胞增殖。功能分析表明， 促进细胞分化，并在成肌细胞分化过程中增强肌球蛋白重链（Myosin heavy chain，MyHC）和肌细胞增强因子 2（Myocyte enhancer factor 2，MEF2）的表达。通过下拉和 RNA 免疫沉淀（RNA Immunoprecipitation，RIP）实验，发现 lncRNA 与 PBX/同源盒 2（PBX/Knotted Homeobox 2，PKNOX2）相互作用。CUT 和 Tag 实验表明，lncRNA 敲低后，PKNOX2 显著富集在 启动子上。我们的研究结果表明，lncRNA 与 PKNOX2 的相互作用解除了 PKNOX2 对 启动子的抑制作用，增加了其表达，并促进了 PSC 的分化。该 lncRNA 的新作用揭示了 lncRNA 调节猪肌发生的分子机制。
